This is a revised application which is focused on the analysis of the mechanism of dRpase activity in bacteria, and the identification of corresponding activities in yeast. The specific aims are: (1) A novel plasmid assay construct will be used to analyze the role of the several known dRpase activities in bacterial DNA base excision repair. The plasmid assay construct will also be used to determine if the repair process involves replacement of one or several nucleotides. (2) Interactions between several proteins that function as part of the DNA base excision repair pathway will be investigated by use of the yeast two- hybrid system, and where appropriate by oligo-histidine affinity chromatography. Interactions will also be studied with purified repair enzymes and the plasmic assay system of SA1, in vitro, by analysis of repair rates as a function of added protein co-factors. (3) Characterization and purification of a eukaryotic dRpase activity in the yeast S. pombe. The yeast enzyme will be purified and subsequently subjected to peptide sequencing so as to yield oligonucleotide probes which can be used to screen a yeast cDNA library for the corresponding mRNA. It is proposed that this would allow for the sequencing, cloning, and possible source to identify a homologous human allele.